The
modulatory role of M2 muscarinic receptor on
apomorphine-induced yawning and genital
grooming
Gamberini MT, Bolognesi ML, Nasello AG.
Department of Physiological
Sciences of the Santa Casa de São Paulo -
Faculty of Medical Sciences, Brazil.
Abstract
The interaction between dopaminergic and
cholinergic pathways in the induction of
behavioral responses has been previously
established. In the brain, M2 receptors are
found predominantly in presynaptic cholinergic
neurons as autoreceptors, and in dopaminergic
neurons as heteroceptors, suggesting a control
role of acetylcholine and dopamine release,
respectively.
Our aim was to investigate the role of M2
receptors on the yawning and genital grooming of
rats induced by apomorphine, a dopaminergic
receptor agonist, focusing on the interaction
between cholinergic and dopaminergic pathways.
Initially, the effect of atropine, a
non-selective muscarinic antagonist, on yawning
and genital grooming induced by apomorphine
(100?g/kg s.c.) was analyzed. Atropine doses of
0.5, 1 and 2mg/kg i.p. were administered to
Wistar rats 30min before induction of the
behavioral responses by apomorphine.
Number of yawns and time spent genital
grooming were quantified over a 60min period.
Apomorphine-induced yawning was increased by low
dose (0.5mg/kg i.p.) but not by high doses (1
and 2mg/kg, i.p.) of atropine. Genital grooming
was antagonized by 2mg/kg i.p. of atropine and
showed no changes at the other doses tested.
Tripitramine, a selective M2 cholinergic
antagonist, was used as a tool for
distinguishing between M2 and all other
muscarinic receptor subtypes in yawning and
genital grooming. Tripitramine doses of 0.01,
0.02 and 0.04?mol/kg i.p. were administered to
Wistar rats 30min before apomorphine (100?g/kg
s.c.). Number of yawns and time spent genital
grooming were also quantified over a 60min
period. Tripitramine 0.01?mol/kg increased all
parameters. Higher doses, which possibly block
all subtypes of muscarinic receptor, did not
modify the response of apomorphine, suggesting a
non-selective effect of tripitramine at these
doses.
Given that low doses of tripitramine
increased the behavioral responses induced by
apomorphine and that the main distribution of
the M2 receptor is presynaptic, we raised the
hypothesis that tripitramine may alter
cholinergic and/or dopaminergic transmission in
brain areas responsible for induction of yawning
and genital grooming in rats, possibly by
control of acetylcholine and/or dopamine
release.
In addition, the present study showed the
involvement of M2 cholinergic receptors in the
complex mechanisms of functional interactions
between dopaminergic and cholinergic systems
involved in the control of yawning and genital
grooming.
Gamberini
MT, Bolognesi ML, Nasello AG. The modulatory
role of M2 muscarinic receptor on
apomorphine-induced yawning and genital
grooming. Neuroscience Letters 2012;
531:91-95
-Gamberini
MT, Gamberini MC, Nasello AG. Involvement of
dopaminergic and cholinergic pathways in the
induction of yawning and genital grooming by the
aqueous extract of Saccharum officinarum L.
(sugarcane) in rats. Neuroscience Letters
2015;584:270-275
-Naselo
A, Tieppo C, Felicio L Apomorphine induced
yawning in the rat : influence of fasting and
time of day Physiology & Behavior
1995;57(5):967-971
-Nasello AG
et al Modulation by sudden darkness of
apomorphine-induced behavioral responses
Physiology & Behavior 2003;78:521-525
1. Introduction
Dopaminergic and cholinergic transmissions
are involved in 29 motor and cognitive
physiological processes. Imbalances between 30
these transmissions are found in some disorders,
such as Parkin- 31 son's disease, Huntington's
disease and schizophrenia [19]. The 32
pharmacological strategies used for the
treatment of these diseases 33 are based on
restoring the dopamine (DA)/acetylcholine (ACh)
balance in the CNS.
In rats, the interrelationship between
dopaminergic and cholinergic transmission can be
explored through behavioral tools such as
yawning and genital grooming. The involvement of
the dopaminergic system in the induction of
yawning and genital grooming becomes evident by
the administration of small doses of
apomorphine, a direct DA receptor agonist, with
marked affinity for DA D2-like (D2/D3/D4)
receptors.
Results of microinjection studies suggest
that the induction of yawning by D2-like
agonists may be mediated by their action in the
striatum or septum. Also, the action of
acetylcholine in the control of yawning and
genital grooming has been well characterized by
cholinergic agonists and antagonists that induce
or abolish these behaviors, respectively.
A large body of evidence suggests a central
role of cholinergic neurons as a site of action
for the induction of cholinergic yawning and
genital grooming, as well as a common mediator
of these responses induced by a variety of
pharmacological mechanisms. In the striatum, a
high density of dopaminergic and cholinergic
terminals exists and these neurotransmitters
have potent interactions at multiple levels such
as presynaptic regulation of neurotransmitters
and postsynaptic effects in target cells,
including cholinergic neurons.
Another central nucleus capable of initiate
yawning and genital grooming is the
paraventricular nucleus (PVN). One described
mechanism by which dopamine agonist induces
yawning is the activation of D2 receptors. This
elevates nitric oxide synthase activity in the
cell bodies of paraventricular oxytocinergic
neurons projecting to extra-hypothalamic brain
areas, increasing oxytocin release.
Subsequently, oxytocin activates cholinergic
neurotransmission in the hippocampus and
reticular formation of the brainstem.
Microinjections of oxytocin into the medial
preoptic area were also shown to induce
grooming. All five muscarinic receptor subtypes
(M1&endash;M5) are expressed in the CNS. In situ
hybridization and immunohistochemical studies
have shown that M1 and M4 are present
predominantly in efferent neurons as
heteroreceptors, but also in cholinergic neurons
as autoreceptors. M2 is expressed predominantly
in striatal interneurons as a presynaptic
autoreceptor in cholinergic neurons, and as a
heteroreceptor in dopaminergic neurons. The
suppression of striatal acetylcholine release by
M2/M4 autoreceptors, together with the effects
of M2/M4 action on dopamine transmission, lend
support to the hypothesis of modulation of
cholinergic pathways on dopaminergic
transmission in striatum.
Considering the close relationship between
dopaminergic and cholinergic transmission in
yawning and genital grooming, the aim of this
study was to explore a possible cholinergic
modulation of dopaminergic neurotransmission
using a selective antagonist of muscarinic M2
receptor subtype, tripitramine.
Discussion
Dopamine (DA) and acetylcholine (ACh) have
potent interactions in different areas of the
CNS at multiple levels, including presynaptic
regulation of neurotransmitter release and
postsy-naptic effects in target cells. To
explore potential cholinergic modulation of the
dopaminergic neurotransmission involved in
yawning and genital grooming, we conducted this
study analyzing the behavioral responses of rats
to the antagonists of muscarinic receptor
subtypes.
Our data suggests that cholinergic receptors
are involved in the modulation of the yawning
and genital grooming induced by apomorphine. The
results showed that low doses of atropine (0.5
mg/kg) increased the total number of yawns
induced by apomorphine in rats.
Although the functional interactions between
cholinergic and dopaminergic systems are
complex, it has been hypothesized that
acetylcholine release by M2/M4 autoreceptors,
together with the effects of M2/M4 action on
dopamine transmission, modulates dopaminergic
transmission. Since atropine shows selectivity
at M2 receptors in rats: M4 > M5 > M1 >
M2 > M3, the potentiation in yawning induced
by the lower dose of atropine may be explained
by a possible antagonist effect on M2 receptors,
blocking the negative feedback of acetylcholine
release and/or suppressing inhibition of
dopamine release.
We have also considered the possible
antagonistic effect on other cholinergic
receptors of atropine, as a possible explanation
for the blocking of genital grooming by higher
doses of atropine (2 mg/kg). These results led
to the hypothesis that different doses of
atropine antagonize the muscarinic receptor
subtypes differently.
The M2 cholinergic receptors have been
characterized in a variety of areas of the CNS,
including areas associated to yawning and
genital grooming. Hindbrain, brainstem, and
midbrain regions, such as the cerebellum (75%
muscarinic receptor density, mrd), pons/medulla
(70% mrd), and thalamus/hypothalamus (43% mrd)
are enriched with M2 receptors.
In contrast, forebrain regions contain
markedly lower percentages of M2 receptors, with
the cortex expressing 20%, hippocampus 19% and
olfactory tubercle 20%, of total receptor
density. M2 muscarinic autoreceptors are
predominant in the striatum, but the presence of
postsynaptic receptors has also been described
in the dorsal striatum of rat.
Tripitramine displays a unique binding
profile for muscarinic receptor subtypes: M2
> M4 ³ M1 > M3, distinguishing between M2
and all other muscarinic receptor subtypes.
Therefore, we used tripitramine as a tool for
exploring the specific involvement of M2
cholinergic receptor in yawning and genital
grooming induced by the dopaminergic agonist,
apomorphine. Low doses of tripitramine (0.01
mol/kg) were able to potentiate the behavioral
response induced by apomorphine, increasing the
number of total yawns and total time spent
genital grooming in rats compared to control
animals.
The same effects were not obtained when
tripitramine doses were increased. We speculated
that tripitramine, at high doses, loses its
specificity for M2 cholinergic receptors,
whereas it may also antagonize M1 cholinergic
receptor, essential for these behavioral
responses. Given that low doses of tripitramine,
a selective M2 cholinergic receptor antagonist,
increased the behavioral responses induced by
apomorphine and that the main distribution of M2
receptor is presynaptic, it was suggested that
the presynaptic cholinergic control of
acetylcholine and/or dopamine release in brain
areas could be involved in the potentiation of
yawning and genital grooming.
It was previously shown that M2 antagonists
can facilitate memory storage by potentiating
cholinergic tone. This potentiation may have
inhibited dopaminergic bursts with resultant
enhanced yawning and genital grooming.
In psychosis, schizophrenia, pounding,
Tourette's syndrome and hyperkinesias in
general, antidopaminergic drugs were used
chronically and some patients displayed various
side-effects.
Accordingly, the treatment of choice for
these complications constitutes cholinergic
drugs. Furthermore, typical and atypical
antipsychotic drugs were shown to increase the
extracellular release of acetylcholine in the
striatum.
Thus, it can be speculated that the
therapeutic effect of antipsychotic drugs may be
partly mediated via a stimulation (or
disinhibition) of striatal acetylcholine
release, which in turn helps normalize the
functional interaction between
acetylcholine&endash;dopamine systems
particularly in the ventral striatum thereby
restoring a more optimal balance in the activity
of these systems. Hence, cholinergic drug-based
treatments are still used but more specific
effects on the dopaminergic system through M2
antagonism could be investigated to
simultaneously enhance cholinergic transmission
and block dopaminergic transmission.
-Naselo
AG, Tieppo C, Felicio L Apomorphine induced
yawning in the rat: influence of fasting and
time of day Physiology & Behavior
1995;57(5):967-971
-Nasello AG
et al Modulation by sudden darkness of
apomorphine-induced behavioral responses
Physiology & Behavior 2003;78:521-525
